tbuckley/cabot-search huggingface dataset:
P13.04.A REPRODUCIBLE CRISPR-SEQ PIPELINE FOR NGS FUNCTIONAL GENOMICS SCREENINGS FOR GLIOMA CELL LINES
Authors: L Kuhlburger, J Mir Pedrol, L A Haeusser, B Walter, G Gabernet, D Merk, G Tabatabai, S Nahnsen
Journal: Neuro-Oncology (2023)

Abstract BACKGROUND The CRISPR/Cas9 genome editing technology has revolutionized the field of functional genomics by enabling precise editing of DNA. Over the past few years, CRISPR technology has been widely employed to identify genetic dependencies in a broad spectrum of applications paving the way for the discovery and validation of relevant molecular targets, for example potential drug targetsGlioblastoma are incurable primary tumors of the central nervous system. Functional genomics could enable the discovery of novel therapeutic targets or even combination therapies. MATERIAL AND METHODS Here, we present a bioinformatics pipeline that combines functional screens and target editing analysis to identify potential vulnerabilities and drug targets in glioblastoma or other diseases. The pipeline consists of two workflows, one performing functional screening and one performing base editing analysis. This developed CRISPR-seq pipeline is reproducible and containerized, allowing high-throughput processing. All processes are modularized in Nextflow DSL2 (Domain Specific Language) and versioned which facilitates maintainability and customization, as well as the addition of new tools. A quality control report is also automatically generated, allowing the user an overview of their ran experiments. The pipeline also helps us to keep the data FAIR (Findable, Accessible, Interoperable and Reusable) and optimized, making it easier to share with other researchers and collaborate on future studies. RESULTS This pipeline was applied to functional screens using CRISPR/Cas9 genome-wide knockout or activation libraries to detect genetic vulnerabilities that might enhance drug treatment efficacy, e.g. for ataxia telangiectasia and Rad3 related (ATR) inhibition in experimental glioma.With this pipeline, we were able to identify several genes that are involved in drug resistance in glioblastoma, by starting from raw data (fastq files) to genes presenting vulnerabilities.The results obtained through our pipeline provided important information on the disease's molecular mechanisms, which were subsequently confirmed through wet lab validation. CONCLUSION In conclusion, the crisprseq software described in this abstract is a powerful tool for identifying potential genetic dependencies. The pipeline aims at analyzing this output data in a reproducible and containerized way using Nextflow and respecting nf-core community standards. By combining functional screens and base editing analysis with bioinformatics tools, this pipeline allows for the identification of essential genes and pathways, as well as the characterization of specific mutations that contribute to the development and progression of glioblastoma.
P20.03.A PREDICTING THE CLINICAL PHENOTYPE IN<i>NF2</i> RELATED SCHWANNOMATOSIS PATIENTS USING NEW GENETIC SCORES
Authors: R Paterra, A Borghi, N Rosati, M Ibba, D Menicatti, M Moscatelli, I Vetrano, M Saini, V Nazzi, A Silvani, M Eoli
Journal: Neuro-Oncology (2023)

Abstract BACKGROUND NF2 related Schwannomatosis (NF2 -SCHW MIM # 101000) is a rare autosomal dominant familial cancer syndromes caused by mutations in the NF2 gene, mainly due to NF2 point mutations or intragenic deletions. NF2 patients. In 95% of patients, bilateral vestibular schwannomas are present, but NF2 -SCHW patients can develop other tumors and show ophthalmic and dermatological signs. Several studies established a genotype-phenotype correlations; the UK NF2 Reference Group proposed a Genetic Severity Score (GSS) based on the type and location of NF2 germline variant observed. Catasus et al. after validation of the score in a Spanish cohort suggested a new score based also on a functional assay of Merlin and its downstream pathways (FGSS). Recently Teranish et al. used targeted deep sequencing to predict functional prognosis in those patients. MATERIAL AND methods 70 patients (median age 40) were identified by scanning the electronic NF2-SCHW patient database at Fondazione IRCCS C. Besta, where the patients had undergone full neurological, ophthalmic, and audiological assessment. The median follow-up was 14 yrs. Clinical diagnosis was established following the Updated diagnostic criteria and nomenclature for neurofibromatosis type 2 and schwannomatosis”. All patients underwent NF2, SMARCB1 and LZTR1 mutational screening by NGS and MPLA analysis using blood DNA or the tissue of two different tumors when available.NF2 gene germline pathogenic were found in 45 subjects (64%), in most cases NF2 point mutations, but in 4 cases and whole gene deletions was observed and in 2 a ring NF2 chromosome was identified, in four cases a mosaic NF2-SCH was identified and in 21 (30%) patients no pathogenic variant was found. We assessed NF2-SCHW clinical phenotype in relation to the UK GSS and FGSS to validate their use as clinical and research tools, despite the significant phenotypic variability observed. RESULTS The disease outcome differed significantly depending both on clinicaland genetic factors. Among these factors,” Age of symptom onset” “Truncating”, “Mosaic”, and “whole gene deletion” had the most significant effects on functional disability. Conclusion GSS and FGSS showed significant correlation with several measures, allowing stratification of patients with severe and mild disease but not with moderate phenotype. Furthermore, we provided evidence on correlation between whole NF2 deletion extent and phenotype severity. Large cohorts of NF2-SCHW patients are needed to identify more accurate scores, becoming useful tools for patient management.
P10.02.B THE PSYCHOTROPIC AGENT THIORIDAZINE IMPEDES MELANOMA BRAIN METASTASIS CELL ACTIVITY THROUGH INHIBITION OF AUTOPHAGY
Authors: E Moen Taule, T Wikerholmen, K Sarnow, E Rigg, B Feginn Berle, T Sundstrøm, J Wang, F Thorsen
Journal: Neuro-Oncology (2023)

Abstract BACKGROUND Neural tissue and melanocytes share the same germ layer in early embryonic development. It is then perhaps not by coincidence that melanoma is one of the most common cancer types to metastasize to the brain. With aggressive treatment the median survival of melanoma brain metastases (MBM) is now estimated to be approximately 13 months. Targeted therapies and immunotherapy have armed us with new systemic weapons and shown great responses, but unfortunately most of these patient relapse. Drug repurposing, using old drugs for new purposes, has had several successes for other diseases. The anti-psychotic drug thioridazine (THD), a dopamine receptor 2 antagonist, with properties to cross an intact blood brain barrier, has earlier shown promising effects on melanoma cells that escaped prior treatment and on cancer stem cells. However, the therapeutic effects on MBM have not been previously investigated. MATERIAL AND METHODS We used three BRAF V600E (H1, H2 and H10) and one L577F mutated patient derived MBM cell lines (H3) developed in our lab in our experiments. A monolayer viability assay (WST-1) was utilized to determine IC50 doses. Migrational and proliferational effects of THD was studied by IncuCyte live cell imaging. Colony forming properties and the ability of cells to grow in an anchorage independent environment after THD treatment was investigated using a 2D clonogenic assay and a 3D soft agar model. A potential therapeutic window was assessed through normal human astrocytes (NHA) and fetal rat brain organoids (FRBO). Apoptosis and necrosis were studied by Annexin V/propidium iodide (PI) double staining followed by flow cytometric analysis after 72 hours of treatment. Western blots after THD treatment was done to elucidate underlying molecular mechanisms. RESULTS Cell viability studies showed that THD exerted a dose-dependent inhibition of tumor cell viability (IC50 doses between 9-12 µM), while NHA tolerated similar drug concentrations. Tumor cell migration and proliferation were almost completely inhibited with concentrations below the IC50 doses. An inhibition of clonogenic formation and 3D growth was also observed. Bright field microscopy showed changes in MBM morphology in monolayers, but FRBOs were intact after drug treatment. Apoptosis in MBM cells was induced after treatment with THD through PARP/cleaved caspase 3/Bcl-2. Expression levels of LC3B-II and p62 were significant increase in treated cells, indicating inhibition of autophagic flux at a late stage. CONCLUSION Our data shows that THD has a strong selective cytotoxicity in MBM cells through inhibition of autophagic flux, with a minimal effect on NHA and FBRO.
P11.11.A VOLUMETRIC AND SPATIAL ANALYSES OF TREATMENT-RELATED INJURY IN GLIOMA
Authors: A T J van der Boog, S David, M Vreeman, A A M Steennis, F Cialdella, J Dankbaar, T J Snijders, J J C Verhoeff, P A Robe
Journal: Neuro-Oncology (2023)

Abstract BACKGROUND As diffuse glioma cannot yet be cured, benefits of therapeutic strategies should be carefully balanced against their burden. Postoperative ischemia is a relatively common adverse event of surgery, which can lead to serious neurological deficits. Radiation therapy (RT) can cause post-RT lesions, which may cause clinical symptoms and can create a therapeutic impasse since they mimic tumor progression. We aimed to explore the spatial and volumetric tumor and treatment characteristics in relation to development of treatment-related effects in glioma. Materials and METHODS We retrospectively included 144 WHO grade II-IV supratentorial glioma cases, which were resected from 2012 till 2014 and had early postoperative MRI with diffusion-weighted imaging (DWI). Tumors were delineated on preoperative contrast-enhanced T1 MRI for high grade gliomas and FLAIR for low grade gliomas. Postoperative ischemia was defined as new, confluent diffusion restriction on postoperative DWI. Post-RT injury was investigated in patients who received RT after first surgery and had at least 6 months of follow-up (FU), and was defined as new or increasing contrast-enhancement that subsequently stabilized or decreased during FU. After manual delineation and non-linear registration to MNI stereotaxic space, all lesions were investigated with regard to size and location of pre-operative tumor, resection cavity, residual tumor and extent of resection (EOR). Voxel-based analysis (VBA) was performed with family-wise error correction for multiple comparisons. RESULTS Postoperative ischemia occurred in 93 of 144 cases and was associated with a larger EOR (P=0.02). Ischemic volume was correlated with a larger preoperative tumor volume (P=0.009) and resection volume (P=0.001). Post-RT injury was found in 25 of 67 analyzed cases and was associated with a larger residual tumor volume (P=0.021) and a smaller EOR (P=0.005). VBA showed an association of the occurrence of post-RT lesions with a cluster of preoperative tumor voxels in the left temporal lobe and clusters of residual tumor voxels in the left temporal and right frontal region. No association in volume or location was found between postoperative ischemia and post-RT lesions. CONCLUSION Adverse events of glioma treatment may be influenced by tumor and treatment volumes. In our population, a larger EOR was associated with the occurrence of postoperative ischemia, and larger tumor and resection volumes were correlated with the volume of ischemia. Additionally, the occurrence of post-RT injury was associated with a larger residual tumor and a smaller EOR, as well as a left temporal preoperative tumor location and a right frontal or left temporal residual tumor location. A thorough examination of spatial and volumetric characteristics of the tumor, treatment and subsequent complications can help elucidate the etiology and guide treatment decisions.
P11.74.A EXPERIENCE OF PREOPERATIVE GAMMA KNIFE RADIOSURGERY FOR RECURRENT BRAIN METASTASES
Authors: E Verhaak, T van Seeters, H Ardon, B Brouwers, P Hanssens
Journal: Neuro-Oncology (2023)

Abstract BACKGROUND Resection of brain metastases (BM) without additional radiation therapy yields a high local failure rate. Surgical resection of newly diagnosed BM combined with post-operative stereotactic radiosurgery (SRS) to reduce the risk of a local recurrence is an accepted and effective treatment option. Drawbacks of postoperative SRS include uncertainty in target delineation, potential delay in the administration of SRS and intraoperative risk of tumor spillage. Preoperative SRS might address these potential drawbacks. We present our experience with preoperative Gamma Knife radiosurgery (GKRS) for recurrent BM. MATERIAL AND METHODS Data of patients with recurrent BM treated with GKRS followed by surgical resection between June 2019 and June 2021 at the Elisabeth-TweeSteden Hospital Tilburg were retrospectively collected. Surgery was necessary according to the multidisciplinary team because of mass effect, a symptomatic lesion or a large tumor volume not eligible for salvage stereotactic radiosurgery. Pre-operative SRS was performed with GKRS followed by surgery within 24 hours. All patients had follow-up appointments with MRI scan as long as clinical meaningful. In case of new intracranial disease new treatment was offered if appropriate. Descriptive analyses were used to give an overview of the patient characteristics. Kaplan-Meier curves were used to analyze overall survival. RESULTS 25 patients (male 8, female 17; median age 64 years (range 20-79 years)) underwent preoperative GKRS for recurrent brain metastases. Most patients were previously treated with GKRS (76%) or SRS (24%). Most patients had non-small cell lung cancer (44%), followed by breast (12%), small cell lung cancer (12%), and melanoma (12%). The median overall survival was 18.5 months (95% CI, 4.9 to 32.1 months). Eight patients (32%) had (multiple) surgical complications, three of these patients died due to the postoperative complications. Twelve patients (48%) developed a local recurrence. The median time to local recurrence was 6.9 months (95% CI, 4.7 to 9.1months). Of the 16 patients with a subtotal resection, 10 patients developed a local recurrence, with 6 recurrences at the site of the macroscopic residual tumor. Two patients (8%) developed leptomeningeal disease at 2.8 months and 3.9 months. Two patients (8%) developed new brain metastases (distant failure) at 5.4 months and 23.8 months. CONCLUSION Pre-operative radiosurgery was well tolerated in a group of patients with recurrent BM who were eligible for surgery. Larger series are needed to perform multivariate analyses on predictors of local recurrence in order to evaluate for which patients this treatment option is best suited.
P13.10.A PREDICTING SURVIVAL OF GLIOBLASTOMAAFTER RADIOTHERAPY USING DEEP LEARNING AND NEUROIMAGING IN A MULTI-CENTRE COHORT
Authors: A Chelliah, D A Wood, L S Canas, H Shuaib, C Linares, A Bassiouny, A Luis, T Young, A Roman, C Dragos, M MacDonald, Y H Lau, S Currie, K Fatania, R Frood, R Mathew, J Brock, E Chandy, S Tenant, C Rowland-Hill, M Williams, Q Wang, E Beaumont, T Lam, L Welsh, K Foweraker, J Lewis, S Thust, S Wastling, J Glendenning, L Brazil, A Swampillai, S Ourselin, M Modat, T C Booth
Journal: Neuro-Oncology (2023)

Abstract BACKGROUND Glioblastoma is an aggressive brain tumour routinely monitored with MRI. However, the potential to use imaging as a prognostic biomarker predicting survival after treatment starts is unclear. This study aims to predict survival within eight months of completing radiotherapy from MRIs using deep learning. MATERIAL AND METHODS The dataset consists of 206 prospective and retrospective consecutive patients with glioblastoma (WHO 2021) across 11 UK centres (short-term survival: N=64; 31.1%). Models were trained/validated on N=158 (76.7%) retrospective patients (3 centres). Two holdout sets were sampled: retrospective test set (N=19); and external, prospective test set (N=29; 8 separate centres). Dense convolutional neural networks were developed with separate branches for T1c-w and T2-w MR sequences; a third branch concatenates outputs from those branches to predict survival. Dense blocks used pretrained weights for an abnormality detection model. A semi-supervised approach was implemented; patients without known outcomes were added with pseudo-labels during training. Parameter tuning included the number of updated blocks, linear layer sizes, and weighting of pseudo-labels. Imaging models were compared to counterparts that apply non-imaging variables (demographics, MGMT status/%, treatment). Logistic regression, support vector classifier, and decision tree models were fit on non-imaging features alone with feature selection and mean/mode imputation. Combined imaging/non-imaging networks were developed that added a branch for non-imaging inputs. RESULTS The current best-performing imaging model used MR sequences alone (validation weighted-ROC=0.80, 95% CI=0.75-0.84; balanced accuracy=0.70, 95% CI=0.55-0.85). Among non-imaging models, the logistic regression model with feature selection performed best (validation weighted-ROC=0.59, 95% CI=0.56-0.63; balanced accuracy=0.59, 95% CI=0.56-0.63). Combined model development is ongoing. CONCLUSION This is the first known model applying deep learning to MRIs from multiple centres to distinguish patients with failure to respond to chemoradiotherapy, and those who survive the subsequent treatment window. Based on initial results, image-based models performed best at predicting survival. Such models could be used across centres to suggest closer monitoring and trial targeting of patients with expected short-term survival compared to treatment response.
P15.13.B DEVELOPMENT OF [18F]AG-120 AS AN IMAGING AGENT FOR THE DETECTION OF THE MUTANT ISOCITRATE DEHYDROGENASE 1 IN GLIOMA BY PET
Authors: M Toussaint, T Lai, S Dukic-Stefanovic, R Teodoro, R Moldovan, S Meister, M Ullrich, J Pietzsch, K Kopka, T Juratli, B Wenzel, W Deuther-Conrad
Journal: Neuro-Oncology (2023)

Abstract BACKGROUND Gliomas are highly invasive and infiltrative solid tumors that present significant clinical challenges in terms of achieving complete surgical resection. These tumors often harbor mutated isocitrate dehydrogenase enzymes (mIDH), making them attractive theranostic targets. Therefore, accurately assessing mIDH status is essential for effective patient management. Currently, the presence of mIDH1 is determined either by invasive biopsy or indirectly by measuring the mIDH-derived oncometabolite 2-hydroxyglutarate (2-HG) using magnetic resonance spectroscopy (MRS). The availability of mIDH-targeted radioligands would allow for PET neuroimaging to detect the mIDH1 protein directly and non-invasively. MATERIAL AND METHODS The study involved obtaining 18F-labeled AG-120 (Ivosidenib), an FDA-approved small molecule inhibitor of mIDH, by Cu-mediated radiofluorination of a corresponding diastereomerically pure stannyl precursor. The researchers studied the internalization of [18F]AG-120 in vitro in U251 human glioblastoma cells stably transfected with IDH1 or IDH1R132H. Brain uptake and metabolism of [18F]AG-120 were investigated in healthy CD-1 mice by dynamic PET-MR imaging and radio-chromatographic analyses of plasma and brain tissue. Dynamic PET-CT imaging studies were performed in nude rats bearing U251-IDH1 or U251-IDH1R132H glioblastoma. RESULTS The (S,S)-enantiomer of [18F]AG-120 was successfully prepared, and the procedure was implemented into a remotely-controlled radiosynthesis module. A considerably higher uptake of activity was observed in U251-IDH1R132H cells compared to U251-IDH1 cells (0.422 vs. 0.014% applied activity/μg protein after 120 min incubation). The percentage of non-metabolized [18F]AG-120 in the mouse 30 min post i.v. injection was 85% in plasma and 91% in the brain, respectively. Dynamic PET studies showed limited blood-brain barrier permeation of [18F]AG-120 (SUV &amp;lt; 0.1 within the first 5 min post i.v. injection), which could be significantly increased by inhibiting the p-glycoprotein 1 with cyclosporine A. PET studies with glioblastoma rats indicated slightly higher tumor-to-blood ratios in IDH1R132H-tumors compared to IDH1-tumors (~1.7 vs. ~1.3 at 40-60 min p.i.). CONCLUSION For the first time, pure (S,S)-[18F]AG-120 has been synthesized. In addition, GMP-compliant production is facilitated by the established automated radiosynthesis. Despite limited BBB permeation, [18F]AG-120 shows target-specific internalization in vitro and high metabolic stability in vivo. Furthermore, the slightly higher accumulation of [18F]AG-120 in IDH1R132H-glioblastoma compared to IDH1-glioblastoma in vivo encourages further evaluation of mIDH inhibitors for the noninvasive detection of IDH1R132H in glioma by PET. SUPPORT European-Regional-Development-Fund ERDF and Sächsische-Aufbaubank SAB (#100364142).
P20.07.A IDENTIFYING SELF-EFFICACY DIFFERENCES IN ADULT RARE CNS PATIENTS PARTICIPATING IN A NOVEL WEB-BASED NATURAL HISTORY
Authors: A Acquaye-Mallory, E Vera, A Choi, K Wall, T Kunst, M R Gilbert, T S Armstrong
Journal: Neuro-Oncology (2023)

Abstract BACKGROUND A cancer diagnosis can diminish self-efficacy (sense of control and confidence), which can be essential for self-management. Perceived control in coping effectively with the adverse effects of a diagnosis has been correlated with improved mood, self-regulation, and decreased symptom distress. Using data collected in our global web-based NCI-CONNECT Natural History study (https://www.cancer.gov/rare-brain-spine-tumor/ ), we explored the relationship between self-efficacy levels in managing social interactions (SEMSI), sample characteristics, and mood-related inference (REM) in adult patients with rare CNS tumors. MATERIAL AND METHODS Interested adult patients completed a study enrollment confirming a rare diagnosis. Eligibility and consent forms were reviewed with the study coordinator before consent was signed. A unique link to complete the online survey was sent to enrolled patients. This web-based survey captured current demographic/clinical data, symptom burden, and REM (mood, relations with others, enjoyment of life, scores ≥ 2=moderate-severe (M-S)) from the MDASI-BT &amp; MDASI-SP and the PROMIS SEMSI (t-scores: Low=&amp;lt;40 (lower confidence), Average=50 (US general population), High= &amp;gt;60 (greater confidence)). RESULTS Evaluated patients (n=166) were predominantly female (72%), the median age was 45 (19-75, and 15% were on active treatment. An ependymoma diagnosis (66%), no recurrence (80%), and tumor location in the brain (53%) were common. Median time since diagnosis was 14 months (0-402 months). Most patients reported average SEMSI, none reported high, and 28% of brain vs. 7% of spine reported low SEMSI. Among demographic/clinical factors, low SEMSI correlated with lower education (r=0.22) and a prolonged time between symptoms and surgery (r= -0.29) among brain patients. Additionally, married (42%) brain patients had a higher chance of endorsing low SEMSI. No associations were identified for spine patients. Brain patients, reporting low (92%) and average (48%) SEMSI, also endorsed M-S REM (p&amp;lt;0.001, OR=12.1, n=87). Additionally, all M-S REM symptoms (fatigue, disturbed sleep, distress, sadness, irritability) were associated with low SEMSI. As REM worsened, SEMSI decreased in brain patients (r= -0.54), with no relationship found in spine patients. CONCLUSION We found that brain tumor patients reported less confidence in managing social interactions with low self-efficacy levels linked to marital status, education, a longer time between symptoms and surgery, and worsened mood-related interference, but surprisingly no association in those with spine tumors. Identifying patient characteristics associated with self-efficacy within tumor locations can lead to targeted cognitive and behavioral interventions motivating patients to mitigate distress that can help with adjustment to one’s disease.